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Interleukin(IL)-36α and IL-36γ Induce Proinflammatory Mediators from Human Colonic Subepithelial Myofibroblasts.

Kanda T, Nishida A, Takahashi K, Hidaka K, Imaeda H, Inatomi O, Bamba S, Sugimoto M, Andoh A - Front Med (Lausanne) (2015)

Bottom Line: The mRNA expression of proinflammatory mediators induced by IL-36α and/or IL-36γ was significantly suppressed by transfection of siRNA for MyD88 or IRAK1.Both inhibitors of mitogen activated protein kinases and siRNAs specific for NF-κBp65 significantly reduced the expression of IL-6 and CXC chemokines induced by IL-36α and/or IL-36γ.These results suggest that IL-36α and IL-36γ contribute to gut inflammation through the induction of proinflammatory mediators.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Shiga University of Medical Science , Otsu , Japan.

ABSTRACT

Background: Interleukin (IL)-36 cytokines are recently reported member of the IL-1 cytokine family. However, there is little information regarding the association between IL-36 cytokines and gut inflammation. In the present study, we investigated the biological activity of IL-36α and IL-36γ using human colonic subepithelial myofibroblasts (SEMFs).

Methods: The mRNA expression and the protein expression of target molecules in SEMFs were evaluated using real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. The intracellular signaling of IL-36 cytokines was analyzed using Western blot analysis and small interfering RNAs (siRNAs) specific for MyD88 adaptor proteins (MyD88 and IRAK1) and NF-κB p65.

Results: IL-36α and IL-36γ significantly enhanced the secretion of IL-6 and CXC chemokines (CXCL1, CXCL2, and CXCL8) by SEMFs. The combination of IL-36α/γ and IL-17A or of IL-36α/γ and tumor necrosis factor-α showed a synergistic effect on the induction of IL-6 and CXC chemokines. The mRNA expression of proinflammatory mediators induced by IL-36α and/or IL-36γ was significantly suppressed by transfection of siRNA for MyD88 or IRAK1. Both inhibitors of mitogen activated protein kinases and siRNAs specific for NF-κBp65 significantly reduced the expression of IL-6 and CXC chemokines induced by IL-36α and/or IL-36γ.

Conclusion: These results suggest that IL-36α and IL-36γ contribute to gut inflammation through the induction of proinflammatory mediators.

No MeSH data available.


Related in: MedlinePlus

Role of MyD88 adaptor proteins (MyD88 and IRAK1) in IL-36 signaling in human colonic SEMFs. SEMFs were transfected with siRNA specific for the MyD88 adaptor proteins (MyD88 and IRAK1) or with control siRNA. The cells were incubated for 24 h with or without 100 ng/ml of IL-36α (upper panels) or IL-36γ (lower panels). The mRNA expression of IL-6 and CXC chemokines (CXCL1, CXCL2, and CXCL8) was then evaluated using real-time PCR. The mRNA expression for IL-6 and chemokine was converted to a value relative to β-actin mRNA expression and presented as fold-increase relative to the results for medium alone (no stimulation). Data are expressed as means ± SD of four independent experiments. *P < 0.05, **P < 0.01; significant differences from the values for IL-36α or IL-36γ stimulation.
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Figure 4: Role of MyD88 adaptor proteins (MyD88 and IRAK1) in IL-36 signaling in human colonic SEMFs. SEMFs were transfected with siRNA specific for the MyD88 adaptor proteins (MyD88 and IRAK1) or with control siRNA. The cells were incubated for 24 h with or without 100 ng/ml of IL-36α (upper panels) or IL-36γ (lower panels). The mRNA expression of IL-6 and CXC chemokines (CXCL1, CXCL2, and CXCL8) was then evaluated using real-time PCR. The mRNA expression for IL-6 and chemokine was converted to a value relative to β-actin mRNA expression and presented as fold-increase relative to the results for medium alone (no stimulation). Data are expressed as means ± SD of four independent experiments. *P < 0.05, **P < 0.01; significant differences from the values for IL-36α or IL-36γ stimulation.

Mentions: We next investigated whether MyD88 adaptor proteins were involved in the signal transduction of IL-36α or IL-36γ. For this purpose, we employed siRNA transfection system to silence the gene expression of MyD88 adaptor proteins. As shown in Figure 4, siRNA specific for MyD88 or IRAK1, but not control siRNA, significantly suppressed the mRNA expression of IL-6 and CXC chemokines (CXCL1, CXCL2, and CXCL8) in response to IL-36α or IL-36γ [efficacy of siRNAs specific for MyD88, IRAK1, and NF-κB65 was confirmed by real-time PCR (Figure S2 in Supplementary Material)]. These findings indicated that MyD88 adaptor proteins are essential for the intracellular signaling of both IL-36α and IL-36γ for the induction of proinflammatory mediators.


Interleukin(IL)-36α and IL-36γ Induce Proinflammatory Mediators from Human Colonic Subepithelial Myofibroblasts.

Kanda T, Nishida A, Takahashi K, Hidaka K, Imaeda H, Inatomi O, Bamba S, Sugimoto M, Andoh A - Front Med (Lausanne) (2015)

Role of MyD88 adaptor proteins (MyD88 and IRAK1) in IL-36 signaling in human colonic SEMFs. SEMFs were transfected with siRNA specific for the MyD88 adaptor proteins (MyD88 and IRAK1) or with control siRNA. The cells were incubated for 24 h with or without 100 ng/ml of IL-36α (upper panels) or IL-36γ (lower panels). The mRNA expression of IL-6 and CXC chemokines (CXCL1, CXCL2, and CXCL8) was then evaluated using real-time PCR. The mRNA expression for IL-6 and chemokine was converted to a value relative to β-actin mRNA expression and presented as fold-increase relative to the results for medium alone (no stimulation). Data are expressed as means ± SD of four independent experiments. *P < 0.05, **P < 0.01; significant differences from the values for IL-36α or IL-36γ stimulation.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
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Figure 4: Role of MyD88 adaptor proteins (MyD88 and IRAK1) in IL-36 signaling in human colonic SEMFs. SEMFs were transfected with siRNA specific for the MyD88 adaptor proteins (MyD88 and IRAK1) or with control siRNA. The cells were incubated for 24 h with or without 100 ng/ml of IL-36α (upper panels) or IL-36γ (lower panels). The mRNA expression of IL-6 and CXC chemokines (CXCL1, CXCL2, and CXCL8) was then evaluated using real-time PCR. The mRNA expression for IL-6 and chemokine was converted to a value relative to β-actin mRNA expression and presented as fold-increase relative to the results for medium alone (no stimulation). Data are expressed as means ± SD of four independent experiments. *P < 0.05, **P < 0.01; significant differences from the values for IL-36α or IL-36γ stimulation.
Mentions: We next investigated whether MyD88 adaptor proteins were involved in the signal transduction of IL-36α or IL-36γ. For this purpose, we employed siRNA transfection system to silence the gene expression of MyD88 adaptor proteins. As shown in Figure 4, siRNA specific for MyD88 or IRAK1, but not control siRNA, significantly suppressed the mRNA expression of IL-6 and CXC chemokines (CXCL1, CXCL2, and CXCL8) in response to IL-36α or IL-36γ [efficacy of siRNAs specific for MyD88, IRAK1, and NF-κB65 was confirmed by real-time PCR (Figure S2 in Supplementary Material)]. These findings indicated that MyD88 adaptor proteins are essential for the intracellular signaling of both IL-36α and IL-36γ for the induction of proinflammatory mediators.

Bottom Line: The mRNA expression of proinflammatory mediators induced by IL-36α and/or IL-36γ was significantly suppressed by transfection of siRNA for MyD88 or IRAK1.Both inhibitors of mitogen activated protein kinases and siRNAs specific for NF-κBp65 significantly reduced the expression of IL-6 and CXC chemokines induced by IL-36α and/or IL-36γ.These results suggest that IL-36α and IL-36γ contribute to gut inflammation through the induction of proinflammatory mediators.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Shiga University of Medical Science , Otsu , Japan.

ABSTRACT

Background: Interleukin (IL)-36 cytokines are recently reported member of the IL-1 cytokine family. However, there is little information regarding the association between IL-36 cytokines and gut inflammation. In the present study, we investigated the biological activity of IL-36α and IL-36γ using human colonic subepithelial myofibroblasts (SEMFs).

Methods: The mRNA expression and the protein expression of target molecules in SEMFs were evaluated using real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. The intracellular signaling of IL-36 cytokines was analyzed using Western blot analysis and small interfering RNAs (siRNAs) specific for MyD88 adaptor proteins (MyD88 and IRAK1) and NF-κB p65.

Results: IL-36α and IL-36γ significantly enhanced the secretion of IL-6 and CXC chemokines (CXCL1, CXCL2, and CXCL8) by SEMFs. The combination of IL-36α/γ and IL-17A or of IL-36α/γ and tumor necrosis factor-α showed a synergistic effect on the induction of IL-6 and CXC chemokines. The mRNA expression of proinflammatory mediators induced by IL-36α and/or IL-36γ was significantly suppressed by transfection of siRNA for MyD88 or IRAK1. Both inhibitors of mitogen activated protein kinases and siRNAs specific for NF-κBp65 significantly reduced the expression of IL-6 and CXC chemokines induced by IL-36α and/or IL-36γ.

Conclusion: These results suggest that IL-36α and IL-36γ contribute to gut inflammation through the induction of proinflammatory mediators.

No MeSH data available.


Related in: MedlinePlus